Genes are regulated in an inducible, tissue specific or constitutive manner. There are different types of structural elements which are involved in the regulation of gene expression. Cis-acting elements, located in the proximity of, or within genes, serve to bind sequence-specific DNA binding proteins, i.e., trans-acting factors. The binding of proteins to DNA is responsible for the initiation, maintenance, or down-regulation of gene transcription.
Cis-acting elements which control genes include promoters, enhancers and silencers. Promoters are positioned next to the transcription start site and function in an orientation-dependent manner, while enhancer and silencer elements, which modulate the activity of promoters, may be flexible with respect to their orientation and distance from the transcription start site.
Various promoter sequences are available which may be used in the genetic engineering of plants. Such promoters may be utilized to initiate transcription of a nucleic acid sequence of interest operably linked at the 3' end of the promoter region. Promoters often have transcription specific characteristics such as strength, tissue specificity, developmental stage specificity, etc.
Gene expression in plants may be driven by a number of promoters. Although the endogenous promoter of a gene of interest may be utilized for transcriptional regulation of the gene, the promoter may also be a foreign regulatory sequence. Examples of viral promoters utilized in plant expression vectors include the 35S RNA and 19S RNA promoters of CaMV (Brisson, et al., Nature, 310:511, 1984; Odell, et al., Nature, 313:810, 1985); the full-length transcript promoter from Figwort Mosaic Virs (FMV) (Gowda, et al., J Cell Biochem., 13D: 301, 1989) and the coat protein promoter of TMV (Takamatsu, et al., EMBO J 6:307, 1987). Plant promoters also include the light-inducible promoter from the small subunit of ribulose bis-phosphate carboxylase (ssRUBISCO) (Coruzzi, et al., EMBO J, 3:1671, 1984; Broglie, et al., Science, 224:838, 1984); mannopine synthase promoter (Velten, et al., EMBO J, 3:2723, 1984) nopaline synthase (NOS) and octopine synthase (OCS) promoters (carried on tumor-inducing plasmids of Agrobacterium tumefaciens) and heat shock promoters, e.g., soybean hsp17.5-E or hsp17.3-B (Gurley, et al., Mol. Cell. Biol., 6:559, 1986; Severin, et al., Plant Mol. Biol., 15:827, 1990).
Promoters utilized in plant genetic engineering include both constitutive and inducible natural promoters as well as engineered promoters. The CaMV promoters are examples of constitutive promoters. To be useful, an inducible promoter should 1) provide low expression in the absence of the inducer; 2) provide high expression in the presence of the inducer; 3) use an induction scheme that does not interfere with the normal physiology of the plant; and 4) have no effect on the expression of other genes.
Examples of inducible promoters useful in plants include those induced by chemical means, such as the yeast metallothionein promoter which is activated by copper ions (Mett, et al., Proc. Natl. Acad. Sci., U.S.A., 90:4567, 1993); In2-1 and In2-2 regulator sequences which are activated by substituted benzenesulfonamides, e.g., herbicide safeners (Hershey, et al, Plant Mol. Biol., 17:679, 1991); and the GRE regulatory sequences which are induced by glucocorticoids (Schena, et al., Proc. Natl. Acad. Sci., U.S.A., 88:10421, 1991).
Tissue specific promoters may also be utilized for expression of genes in plants. Tissue specific promoters useful in transgenic plants include the cdc2a promoter and cyc07 promoter (Ito, et al., Plant Mol. Biol., 24:863, 1994; Martinez, et al, Proc. Natl. Acad. Sci. USA, 89:7360, 1992; Medford, et al., Plant Cell, 3:359, 1991; Terada, et al., Plant Journal, 3:241, 1993; Wissenbach, et al., Plant Journal, 4:411, 1993). Additional tissue specific promoters that are utilized in plants include the histone promoter (Atanassova, et al., Plant Journal, 2:291, 1992); the cinnamyl alcohol dehydrogenase (CAD) promoter (Feuillet, et al., Plant Mol. Biol., 27:651, 1995); the mustard CHS 1 promoter (Kaiser, et al., Plant Mol. Biol., 28:231, 1995); the bean grp 1.8 promoter (Keller, et al., Plant Mol. Biol., 26:747, 1994); the PAL1 promoter (Ohl, et al., Plant Cell, 2:837, 1990); and the chalcone synthase A promoter (Plant Mol. Biol., 15:95-109, 1990).